The following contains proprietary/privileged information that Stephen Y Chou requests not be released to persons outside the Government, except for purposes of review and evaluation: The goal of the proposed research is to explore a new single-molecule real-time detector, termed "nanogap detector", and the methods to fabricate this detector, that has the potential to directly sequence single bases of an individual DNA molecule at ultra-high speed and ultra-low cost without labeling or copying, as well as other applications in single DNA analysis and manipulation. Fundamentally different from nanopore-based designs, the nanogap detector uniquely combines a single nanochannel with a nanogap detector (formed by a pair of electrodes with a gap). The nanochannel stretches a single DNA strand into a linear chain and the nanogap sensor inside the channel measures the electrical signal transverse to the DNA backbone as it moves through the channel. Recently, we have fabricated these nanogap detectors, observed, for the first time, the electrical signal perpendicular to the backbone of a single DNA flowing in a nanochannel, and demonstrated (in preliminary tests) sub-5 nm spatial resolution. The detection resolution can be greatly improved by reducing the device dimensions and operating the device in the tunneling-current regime, which is known to have atomic resolution, thus giving us one of the best chances to achieve single base resolution. Moreover, the tip of the electrodes can be functionalized with selected molecules for sensitivity to only one DNA base type, and multiple nanogaps inside one nanofluidic channel for parallel measurements. The proposed research has two parallel interconnected focuses: (a) to explore the nanogap detectors for fast, real-time, de novo, DNA analysis and sequencing without copying and (in many cases) without labeling;and (b) to further develop innovative nanofabrication technologies and designs that drastically improve nanogap detector sensitivity and functionalities (to the benefit of other real-time single DNA detectors). The proposed nanogap detectors offer a wide range of possibilities in revolutionary genome sequencing, from measuring DNA length (label-free) and DNA restriction maps (with markers) to potentially our ultimate goal - high speed single base-pair detection (i.e., from low risk but significant, to high risk and high payoff). The proposed research is based on our many years of experience in nanofabrication and nanodevices in combination with our experience in molecular biology for DNA detection, and will be carried out by a single group of multidisciplinary researchers with electrical engineering, chemistry, biology, and nanotechnology expertise. PROJECT HEALTH RELEVANCE The proposed research is to explore a new single-molecule real-time detector, and the methods to fabricate this detector, that has the potential to directly sequence single bases of an individual DNA molecule at ultra-high speed and ultra-low cost, as well as other applications in single DNA analysis and manipulation.